Identification of a cytoplasmic tropomyosin gene linked to two muscle tropomyosin genes in Drosophila.
Identification of a cytoplasmic tropomyosin gene linked to two muscle tropomyosin genes in Drosophila.
复制标题
果蝇中与两个肌肉原肌球蛋白基因连锁的细胞质原肌球蛋白基因的鉴定。
DOI：
10.1073/pnas.80.23.7123
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发表时间：
1983
影响因子：
11.1
通讯作者：
Storti,RV
中科院分区：
文献类型：
--
作者：
Bautch,VL;Storti,RV
果蝇细胞质原肌球蛋白基因已被鉴定，并与先前鉴定的两个果蝇肌肉原肌球蛋白基因位于相同的基因组DNA克隆上。利用亚克隆基因和非肌肉细胞来源的RNA进行阳性杂交选择翻译实验，获得了一个大小（Mr, 31,000）和等电点（5.0）与脊椎动物细胞质原肌球蛋白相似的蛋白。采用改良的纯化脊椎动物细胞质原肌球蛋白的方法，从果蝇Kc细胞系中部分纯化细胞质原肌球蛋白。根据Kc细胞蛋白的大小、等电点和与多克隆脊椎动物抗原肌球蛋白抗体的交叉反应性，在两步结合实验中，Kc细胞蛋白被鉴定为原肌球蛋白的细胞质形式。Kc细胞质原肌球蛋白与3区基因的杂交选择的体外翻译产物在两个维度上同源，并且这两种蛋白在NaDodSO4/尿素/聚丙烯酰胺凝胶中表现出迁移转移，这是脊椎动物原肌球蛋白的特征。细胞质原肌球蛋白基因与果蝇肌肉原肌球蛋白基因在降低严格条件下杂交。这种交叉杂交跨越了每个肌肉原肌球蛋白基因的几个内部限制性内切酶位点，表明三个果蝇原肌球蛋白基因之间具有部分同源性。这些结果表明果蝇原肌球蛋白是由一个紧密联系的差异调节基因家族编码的。
A Drosophila cytoplasmic tropomyosin gene has been identified and is located on the same genomic DNA clone as two Drosophila muscle tropomyosin genes previously identified. A positive hybrid-selection translation assay using the subcloned gene and RNA from non-muscle cell sources yielded a protein with a size (Mr, 31,000) and isoelectric point (5.0) similar to vertebrate cytoplasmic tropomyosin. A modified protocol for the purification of vertebrate cytoplasmic tropomyosin was used to partially purify cytoplasmic tropomyosin from the Drosophila Kc cell line. The Kc cell protein was identified as a cytoplasmic form of tropomyosin on the basis of its size, isoelectric point, and crossreactivity with a polyclonal vertebrate antitropomyosin antibody in a two-step binding assay. The Kc cell cytoplasmic tropomyosin comigrates in two dimensions with the hybrid-selected in vitro translation product of the region 3 gene, and both proteins show a mobility shift in NaDodSO4/urea/polyacrylamide gels that is characteristic of vertebrate tropomyosins. The cytoplasmic tropomyosin gene hybridizes to both Drosophila muscle tropomyosin genes under decreased stringency conditions. This cross-hybridization spans several internal restriction endonuclease sites in each muscle tropomyosin gene and indicates an overall partial homology among the three Drosophila tropomyosin genes. These results show that Drosophila tropomyosins are encoded by a closely linked family of differentially regulated genes.